Recognition of 8-Oxo-2'-deoxyguanosine in DNA Using the Triphosphate of 2'-Deoxycytidine Connecting the 1,3-Diazaphenoxazine Unit, dCdapTP.

DNA is constantly damaged by various external and internal factors. In particular, oxidative damage occurs in a steady state, and 8-oxo-2'-deoxyguanosine (oxodG) is known as the main oxidative damage. OxodG is a strong genotoxic nucleoside and is thought to be involved in the pathogenesis of cancer and neurological diseases. However, a breakthrough method to detect the position of oxodG in DNA has not yet been developed. Therefore, we attempted to develop a novel method to detect oxodG in DNA using artificial nucleosides. Recently, we have succeeded in the recognition of oxodG in DNA by a single nucleotide elongation reaction using nucleoside derivatives based on a purine skeleton with a 1,3-diazaphenoxazine unit. In this study, we developed a new nucleoside derivative with a pyrimidine skeleton in order to further improve the recognition ability and enzymatic reaction efficiency. We, therefore, designed and synthesized 2'-deoxycytidine-1,3-diazaphenoxazine (Cdap) and its triphosphate derivatives. The results showed that it was incorporated into the primer strand relative to the dG template because of its cytidine skeleton, but it was more effective at the complementary position of the oxodG template. These results indicate that the new nucleoside derivative can be considered as one of the new candidates for the detection of oxodG in DNA.

Unusually elongated superior thyroid cornu associated with recurrent cerebral embolisms.

The superior thyroid cornu (STC) is a normal anatomic structure that forms part of the thyroid cartilage. Here, we report a patient with unusually elongated and ossified STC that might result in recurrent cerebral embolisms. During a second endovascular therapy for recurrent middle cerebral artery embolism, a segment with an irregular filling defect was noted in the internal carotid artery (ICA), at the C1 level. This defect was unnoticed during the initial endovascular procedure. Three-dimensional computed tomography angiography performed after the second endovascular procedure revealed an ICA segment located between the STC and C1 with a tortuous course and irregular wall of the ICA. Therefore, we assumed that STC compression of the ICA could have resulted in thrombus formation at the site and consequent cerebral embolism. The STC should be considered a structure responsible for cerebral embolism. Careful evaluation of the entire ICA course is imperative prior to performing an endovascular thrombectomy for acute embolic occlusion of the middle cerebral artery.

Postoperative tegafur-uracil for stage I lung adenocarcinoma: first real-world data with an exploratory subgroup analysis.

PURPOSE: The effect of postoperative tegafur-uracil on overall survival (OS) after resection of stage I adenocarcinoma has been shown in clinical trials. The purpose of this study was to investigate whether findings from randomized trials of adjuvant tegafur-uracil are reproducible in a real-world setting. METHODS: A retrospective cohort study was performed using a multi-institutional database that included all patients who underwent complete resection of pathological stage I adenocarcinoma between 2014 and 2016. Survival outcomes for patients managed with and without tegafur-uracil were analyzed using the Kaplan-Meier method and a Cox proportional hazards model for the whole patient cohort and in a selected cohort based on eligibility criteria of a previous randomized trial. Propensity score matching was used to adjust for confounding effects. RESULTS: After propensity score matching, the hazard ratios for OS were 0.57 (95% confidence interval (CI) 0.29-1.14, P = 0.11) in the whole cohort and 0.69 (95% CI 0.32-1.50, P = 0.35) in the selected cohort. CONCLUSIONS: The effects of tegafur-uracil in this retrospective study appear to be consistent with those found in randomized clinical trials. These effects may be maximized in patients aged from 45 to 75 years.

Cholesterol Crystals in the Retrieved Thrombus by Mechanical Thrombectomy for Cerebral Embolism: A Case Report and Literature Review.

There are only a few case reports in which cholesterol crystals were found in the thrombus retrieved by mechanical thrombectomy for cryptogenic stroke, leading to a definitive diagnosis. We herein report a case of aortogenic embolic stroke diagnosed by the presence of rich cholesterol crystals in the retrieved thrombus and review the previously reported cases. A woman in her 80s was transferred as an emergency due to consciousness disturbance, right conjugate deviation, and severe left hemiparesis. Magnetic resonance imaging showed occlusion of the right middle cerebral artery (MCA) and acute infarction in the territory. The MCA was recanalized by thrombectomy using an aspiration catheter and stent retriever, and the symptoms improved. Although the physiological examination did not detect the embolic source during hospitalization, pathological examination of the thrombus revealed atheroma with numerous cholesterol crystal clefts and intermixing of fibrin. Contrast-enhanced computed tomography performed based on the pathological results showed atheromatous lesions in the aortic arch as the embolic source. As a subsequent treatment, medications of a strong statin and an antiplatelet agent were continued, and the patient had no recurrence. The finding that the retrieved thrombus is a simple atheroma containing cholesterol crystals with poor hemocytes suggests embolism due to plaque rupture. Pathological examination of the thrombus obtained by thrombectomy is one of the useful diagnostic approaches for stroke etiology and the determination of its treatment.

Epidermal growth factor receptor tyrosine kinase inhibitors as first-line treatment for postoperative recurrent EGFR-mutated lung adenocarcinoma: a multi-institutional retrospective study.

OBJECTIVES: The aim of this study was to analyse the long-term survival outcomes and prognostic factors of patients receiving epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs) as first-line treatment for postoperative recurrent EGFR-mutated lung adenocarcinoma. METHODS: Using a multi-institutional database, we performed a retrospective chart review to identify all patients who had undergone complete resection of stage I-III EGFR-mutated lung adenocarcinoma at 11 acute care hospitals between 2009 and 2016 and had received first-line EGFR-TKI treatment for postoperative recurrence. Adverse events, progression-free survival (PFS) and overall survival (OS) were investigated. Survival outcomes were assessed using Kaplan-Meier analysis. Cox proportional hazards models were used to calculate the hazard ratios (HRs) and 95% confidence intervals (CIs) for PFS and OS. RESULTS: The study sample comprised 154 patients with a median age of 69. The total numbers of events were 101 for PFS and 60 for OS. The median PFS and OS were 26.1 and 55.4 months, respectively. In the multivariable analysis, EGFR ex 21 L858R mutation (HR: 1.71, 95% CI: 1.15-2.55) and shorter disease-free intervals (HR: 0.98, 95% CI: 0.96-0.99) were significantly associated with shorter PFS. Age (HR: 1.03, 95% CI: 1.00-1.07), smoking history (HR: 2.31, 95% CI: 1.35-3.94) and pathological N2 disease at the initial surgery (HR: 2.30, 95% CI: 1.32-4.00) were significantly associated with shorter OS. CONCLUSIONS: First-line EGFR-TKI treatment was generally associated with favourable survival outcomes in patients with postoperative recurrent EGFR-mutated lung adenocarcinoma. EGFR ex 21 L858R mutation may be an important prognostic factor for shorter PFS.

Selective Unnatural Base Pairing and Recognition of 2-Hydroxy-2'-deoxyadenosine in DNA Using Pseudo-dC Derivatives.

The formation of unnatural base pairs within duplex DNA would facilitate DNA nanotechnology and biotechnology. Iso-2'-deoxyguanosine (iso-dG) forms base pairs with iso-2'-deoxycytidine, and its use as an unnatural base pair was investigated. Iso-dG is one of the tautomers of 2-hydroxy-2'-deoxyadenosine (2-OH-dA), known as an oxidatively damaged nucleobase, and its selective recognition in DNA plays an important role in the diagnosis and pathogenesis of disease. Therefore, we focused on pseudo-dC (ψdC) as a suitable molecule that recognizes 2-OH-dA in DNA. Since 2-OH-dA shows tautomeric structures in DNA, we designed and used ψdC, which also has a tautomeric structure. We successfully synthesized a ψdC phosphoramidite compound for the synthesis of oligonucleotides (ODNs) as well as its triphosphate derivative (ψdCTP). Tm measurements revealed that ODNs including ψdC showed stable base pair formation with ODNs having 2-OH-dA. In contrast, low Tm values were observed for other bases (dG, dA, dC, and T). The results obtained for the single-nucleotide primer extension reaction revealed that ψdCTP was incorporated into the complementary position of 2-OH-dA in template DNA with high selectivity. In addition, the primer elongation reaction was confirmed to proceed in the presence of dNTPs. The present study reports an artificial nucleic acid that selectively and stably forms unnatural base pairs with 2-OH-dA in DNA.

Multiple-turnover single nucleotide primer extension reactions to detect 8-oxo-2'-deoxyguanosine in DNA.

The identification of the position of 8-oxo-2'-deoxyguanosine (8-oxo-dG) in DNA is important to clarify the pathogenesis of many diseases. We herein developed a purine-1,3-diazaphenoxazine triphosphate (dPdapTP) and described the first example of detecting the presence of 8-oxo-dG by amplifying it several hundred times after the multiple-turnover single nucleotide primer extension reactions.

Vessel invasion as a predictive factor for recurrence after surgery in stage I lung adenocarcinoma.

BACKGROUND: Patients with early-stage lung cancer who underwent R0 resection often encounter disease recurrence, especially during the early phase; thus, it is deemed vital to determine the predictive factors for recurrence after surgery. In this study, we aimed to identify the independent variables associated with recurrence after complete surgical resection of pathological stage I lung adenocarcinoma. METHODS: We retrospectively reviewed the medical records of 169 patients who underwent pulmonary resection for primary lung adenocarcinoma pathological stage I with curative intent lung cancer surgery from 2015 to December 2018 at our institution for information on the recurrence of the disease. RESULTS: Per the multivariate analysis, the presence of micropapillary pattern and vessel invasion were found to be independent predictors of disease recurrence after surgery (odds ratio [OR]: 9.36, 95% confidence interval [CI]: 2.42-36.2, P = 0.0012; and OR: 4.50, 95% CI: 1.52-13.4, P = 0.0068, respectively). Vessel invasion was also found to be an independent predictor of disease recurrence after surgery within a year (OR 11.4, 95% CI 3.08-42.5, P = 0.0003). CONCLUSIONS: The presence of vessel invasion may help in distinguishing patients with the highest risk of early-phase disease recurrence after surgery. Patients with stage I adenocarcinoma with vessel invasion should undergo intensive surveillance after surgery.

Simple and Easy Synthesis of γ-Amido-dNTPs in Water and Their Polymerase Reaction Properties.

γ-Amido-modified 2'-deoxynucleoside triphosphates (dNTPs) and nucleoside triphosphates (NTPs) are becoming increasingly important as biological tools. We herein describe the simple and easy synthesis of γ-amido-dNTPs and -NTPs from commercially available corresponding dNTPs and NTPs in a one-pot reaction using water-soluble carbodiimide and ammonia solution. We examined the effects of synthesized γ-amido-dNTPs on the DNA polymerase reaction. The results obtained showed the incorporation of these derivatives into the DNA primer while maintaining nucleobase selectivity; however, their incorporation efficiency by DNA polymerase was lower than that of dNTP. This is the first study to demonstrate the successful synthesis of four sets of γ-amido-dNTPs and clarify their properties.

Are volume-dependent parameters in positron emission tomography predictive of postoperative recurrence after resection in patients with thymic carcinoma?

This study aimed to investigate the association between the volume-dependent parameters in 18F-fluorodeoxyglucose-positron emission tomography (18F-FDG PET/CT) and a recurrence of thymic carcinoma. A retrospective chart review was performed based on our multi-institutional database to identify patients undergoing PET prior to resection of thymic carcinoma or neuroendocrine carcinoma between 1991 and 2018. The PET parameters (metabolic tumor volume and total lesion glycolysis) were evaluated retrospectively. The relevant factors were extracted and a survival analysis was performed using the Kaplan-Meier method. Sixteen patients were thus deemed to be eligible for analysis. The median follow-up period following resection was 2.65 years (range: 0.96-0.68 years). The recurrence-free survival was significantly longer in patients with a metabolic tumor volume < = 22.755 cm3 and with total lesion glycolysis < = 105.4006 g/mL (p = 0.001 and 0.001, respectively, by a log-rank test). The metabolic tumor volume and total lesion glycolysis may, therefore, be predictive of the postoperative recurrence of thymic carcinoma.

The treatment and survival of patients with postoperative recurrent thymic carcinoma and neuroendocrine carcinoma: a multicenter retrospective study.

PURPOSE: There are few data available on the outcomes of postoperative recurrent thymic carcinoma (TC) and thymic neuroendocrine carcinoma (TNEC). The aim of this study is to evaluate the treatment and survival in patients with recurrent TC and TNEC after undergoing surgical resection. METHODS: A retrospective chart review was performed using our multicenter database to identify patients with a postoperative recurrence of TC and TNEC from 1995 to 2018. The clinicopathological factors were reviewed and the survival outcomes were analyzed. RESULTS: Sixty patients were identified among 152 patients who underwent resection of TC and TNEC. The median follow-up period from the first recurrence was 14.8 months (range 0-144). The 5-year post-recurrence survival was 23% for the whole cohort. According to a univariable analysis, advanced stage [hazard ratio (HR) 2.81, 95% confidence interval (CI) 1.09-9.54], interval between primary surgery and recurrence (HR 0.97, 95% CI 0.95-0.99), any treatment for recurrence (HR: 0.27, 95% CI 0.13-0.58) and chemotherapy for recurrence (HR: 0.46, 95% CI 0.22-0.95) were significant factors related to post-recurrence survival. CONCLUSIONS: Chemotherapy rather than surgery appears to be the mainstay treatment for managing patients with postoperative recurrent TC and TNEC and it may also be considered in multidisciplinary management. Further studies with a larger sample size are required to confirm our findings.

Ligand Design for Specific MHC Class I Molecules on the Cell Surface.

We have validated that ligand peptides designed from antigen peptides could be used for targeting specific major histocompatibility complex class I (MHC-I) molecules on the cell surface. To design the ligand peptides, we used reported antigen peptides for each MHC-I molecule with high binding affinity. From the crystal structure of the peptide/MHC-I complexes, we determined a modifiable residue in the antigen peptides and replaced this residue with a lysine with an ε-amine group modified with functional molecules. The designed ligand peptides successfully bound to cells expressing the corresponding MHC-I molecules via exchange of peptides bound to MHC-I. We demonstrated that the peptide ligands could be used to transport a protein or a liposome to cells expressing the corresponding MHC-I. This strategy may be useful for targeted delivery to cells overexpressing MHC-I, which have been observed in autoimmune diseases.

A case of intrapulmonary lymphangioma surrounded by pulmonary hilar structures.

A 53-year-old man was referred to our hospital for examination of a nodule in the right hilar region observed on a chest roentgenogram. Computed tomography and magnetic resonance imaging revealed a 34 × 32-mm multilocular cystic mass with partial calcification adjacent to the hilar structures in the right upper lung lobe. The mass was resected using video-assisted thoracoscopic surgery and identified as pulmonary cystic lymphangioma through postoperative histopathological analysis. Intrapulmonary lymphangioma is an uncommon benign tumor, with only few reports of surgically resected cases. We describe our surgical findings and recommend surgical resection where possible for suspected intrapulmonary lymphangioma.

Blood retention and antigenicity of polycarboxybetaine-modified liposomes.

Zwitterionic polycarboxybetaines (PCBs) have gained attention as alternative stealth polymers whose liposomal formulation and protein conjugates were reported not to elicit anti-polymer antibodies. Here, we studied the blood retention and antigenicity of liposomes modified with PCBs focusing on their chemical structures and doses. We compared PCBs with either 1 or 3 (PCB1 or PCB3) spacer carbons between the carboxylate and ammonium groups. PCB3-modified liposomes had a short blood retention, whereas PCB1-modified liposomes demonstrated extended blood retention that was somewhat superior to PEGylated liposome. This confirmed the excellent non-fouling nature of PCB1 reported previously. Interestingly, PCB1-liposome as well as PCB3-liposome elicited specific IgMs toward each PCB. The dose-dependent production of specific IgMs to PCB-liposomes was similar to that of PEGylated liposome, i.e., high doses of PCB-liposomes reduced the production of specific IgMs, termed immunological tolerance. These results indicate the importance of investigating the effect of dose to clarify the existence of antigenicity of stealth polymers.

Extended pleurectomy decortication for Masaoka stage IVa thymoma with massive pleural and pericardial dissemination.

We herein report two cases of Masaoka stage IVa thymoma treated by radical resection via thymothymectomy followed by pleurectomy/decortication (PD). Case 1: a 52-year-old man was diagnosed with a type B1 thymoma. Resection of the right lobe of thymus, dissection of left upper mediastinum, and pleurectomy from anterior chest wall to descending aorta were performed via median sternotomy approach. Pericardial resection followed by decortication of the total visceral pleura was then successfully performed via a posterolateral thoracotomy approach. Case 2: a 48-year-old man was diagnosed with type B2 thymoma. Thymothymectomy and extra-pleural dissection except for the right-side diaphragmatic area were achieved via median sternotomy approach. Resection of the visible disseminated lesions of visceral pleura was performed after pleurectomy of the diaphragmatic area via posterolateral thoracotomy approach. Both patients are disease free at 3 years and 2 years and half, respectively. Extended thymothymectomy followed by PD is a candidate approach for surgical management.

Communicating hydrocephalus and coexisting nonenhancing tumor: An ominous sign for patients with neurofibromatosis type 1?

A 26-year-old woman with familial neurofibromatosis type 1 sustained headache that worsened for 1 month. Neuroimaging revealed a mild ventriculomegaly and nonenhancing lesion in the pons. In spite of repeated cerebrospinal fluid examinations and magnetic resonance imaging, the etiology was not determined. The affected pons markedly enlarged in the following 2 months, with extensive leptomeningeal dissemination. Biopsy through hemilaminectomy of the T9 was diagnosed as glioblastoma multiforme. Prompt histologic examination should be performed when patients with familial neurofibromatosis type 1 manifest communicating hydrocephalus coexistent with a nonenhancing tumor.

Chitin digestibility is dependent on feeding behaviors, which determine acidic chitinase mRNA levels in mammalian and poultry stomachs.

Chitin, a polymer of N-acetyl-D-glucosamine (GlcNAc), functions as a major structural component in chitin-containing organism including crustaceans, insects and fungi. Recently, we reported that acidic chitinase (Chia) is highly expressed in mouse, chicken and pig stomach tissues and that it can digest chitin in the respective gastrointestinal tracts (GIT). In this study, we focus on major livestock and domestic animals and show that the levels of Chia mRNA in their stomach tissues are governed by the feeding behavior. Chia mRNA levels were significantly lower in the bovine (herbivores) and dog (carnivores) stomach than those in mouse, pig and chicken (omnivores). Consistent with the mRNA levels, Chia protein was very low in bovine stomach. In addition, the chitinolytic activity of E. coli-expressed bovine and dog Chia enzymes were moderately but significantly lower compared with those of the omnivorous Chia enzymes. Recombinant bovine and dog Chia enzymes can degrade chitin substrates under the artificial GIT conditions. Furthermore, genomes of some herbivorous animals such as rabbit and guinea pig do not contain functional Chia genes. These results indicate that feeding behavior affects Chia expression levels as well as chitinolytic activity of the enzyme, and determines chitin digestibility in the particular animals.

A Multicenter Feasibility Study of EBUS-TBNA for Potentially Operable Non-Small Cell Lung Cancer: The JMTO LC07-02 Study (UMIN000001280).

BACKGROUND: Endobronchial ultrasound-guided transbronchial needle aspiration (EBUS-TBNA) is a standard procedure for the pathological evaluation of the mediastinal nodal (N2) status of lung cancer; however, its feasibility in potentially operable patients with suspicion of minimal N2 disease remains unestablished. PATIENTS AND METHODS: A prospective multicenter study was conducted to assess the feasibility of EBUS-TBNA in this setting. Patients with clinical stage IIIA-N2 non-small cell lung cancer (NSCLC) and mediastinal nodal enlargement on computed tomography (CT) were eligible; patients were ineligible when CT revealed bulky (> 3 cm in the long-axis diameter) N2 or multiple (≥ 3) station N2. If EBUS-TBNA revealed negative results, surgical staging procedures were mandatory. RESULTS: Among 20 eligible patients, EBUS-TBNA provided pathological confirmation of N2 disease (true-positive) in 12 patients. Among 8 patients with negative results with EBUS-TBNA, 4 patients were pathologically diagnosed as having N2 disease with surgical staging procedures (false-negative), and 4 were finally diagnosed as having non-N2 disease with nodal dissection by thoracotomy (true-negative). As a result, the sensitivity of EBUS-TBNA for N2 evaluation (primary endpoint) was 75.0% (95% confidence interval 47.6-92.7%). No grade 3-5 adverse event were documented. CONCLUSION: EBUS-TBNA is potentially safe and useful in the pathological evaluation of N2 status even in potentially operable NSCLC patients with suspicion of minimal N2 disease on preoperative CT.

Phase II study of adjuvant vinorelbine and cisplatin in Japanese patients with completely resected stage II and III non-small cell lung cancer.

PURPOSE: Adjuvant vinorelbine and cisplatin chemotherapy is recognized as a standard regimen for patients with completely resected stage II and III non-small cell lung cancer (NSCLC). However, efficacy of adjuvant chemotherapy in Japanese phase III trials with cisplatin-containing regimen has been controversial, and data are limited on the long-term outcome of adjuvant vinorelbine and cisplatin chemotherapy for NSCLC patients. METHODS: This was a single-arm phase II study in patients with completely resected pathological stage II or III NSCLC, who had not received prior chemotherapy or radiotherapy. Patients received 4 cycles of vinorelbine [25 mg/m(2) of body surface area (BSA)] and cisplatin (40 mg/m(2) of BSA) on days 1 and 8, every 4 weeks. Primary end point was the 3-year relapse-free survival; secondary end points were overall survival and safety. RESULTS: Between December 2006 and January 2011, 60 patients (40 men and 20 women, median age 64 years) were enrolled; all patients were evaluable for survival and safety. Three-year relapse-free survival rate was 55.0 % (95 % confidence interval 42.4-67.6 %). Three- and five-year overall survival rates were 83.3 and 77.8 %, respectively. There were no chemotherapy-related deaths, and adverse effects were acceptable. CONCLUSIONS: Adjuvant vinorelbine and cisplatin chemotherapy was safe and showed a valid relapse-free survival rate. This regimen could be used as a standard regimen and deserves to be a control arm of trials on adjuvant chemotherapy in the Japanese NSCLC patient population.

Intratumoral Wnt2B expression affects tumor proliferation and survival in malignant pleural mesothelioma patients.

Malignant pleural mesothelioma (MPM) is an aggressive thoracic tumor with a poor prognosis. We performed a comprehensive clinical study on the intratumoral expression of Wnt1, Wnt2B and Wnt5A in MPM. One hundred and seven MPM patients were investigated. Immunohistochemistry was performed to evaluate the intratumoral expression of Wnt1, Wnt2B, Wnt5A, survivin and c-Myc, and the Ki-67 proliferation index. The apoptotic index was evaluated by the TUNEL method. Among the 107 MPMs, 23 MPMs (21.5%) were Wnt1-high tumors, 72 MPMs (67.3%) were Wnt2B-high tumors and 54 MPMs (50.5%) were Wnt5A-high tumors. There was no correlation among the levels of Wnt expression. The percentage of Wnt2B-positive tumors was significantly higher compared to that of the other Wnts (p<0.0001). Furthermore, intratumoral Wnt2B expression significantly correlated with the expression of survivin (p<0.001) and c-Myc (p<0.001). Regarding tumor biology, the Ki-67 proliferation index was significantly higher in the Wnt2B-high tumors than in the Wnt2B-low tumors (p=0.0438). In addition, the overall survival was significantly lower in patients with Wnt2B-high tumors than in those with Wnt2B-low tumors (p=0.0238). A Cox multivariate analysis also demonstrated the Wnt2B status to be a significant prognostic factor in MPM patients (p=0.0042). Intratumoral Wnt2B expression was associated with the expression of survivin and c-Myc, tumor proliferation and patient survival in MPM. Wnt2B is a potential molecular target for the treatment of Wnt2B-overexpressing MPMs.